The mechanisms of drug and hormone action of glycogenolysis are being investigated using a single gene, sex-linked variant (carried in I strain mice) that results in a marked deficiency in phosphorylase b kinase (PBK) activity. As assayed in gastrocnemius muscle extracts I strain had 0.3% of the PBK activity found in the control strains C57BL/ST and Ha/ICR (a Swiss- Webster strain). Nevertheless, I strain mouse skeletal muscle contained protein that cross-reacted with an antiserum to rabbit skeletal muscle PBK, indicating the presence of a structurally altered enzyme protein. The mutant PBK has been shown to convert phosphorylase b to a in both the protein-glycogen complex isolated by differential centrifugation of I strain skeletal muscle extracts and in I strain hemidiaphragms. Analyses of catecholamine-stimulated glycogenolysis in isolated hemidiaphragms incubated in Krebs-Ringer bicarbonate buffer showed that both epinephrine and isoproterenol stimulated glycogenolysis in I strain skeletal muscle, but the concentration-response relationship was different from that of C57BL. In both strains, I-propranolol (10 ng/ml), but not d-propranolol (10ng/ml) blocked isoproterenol(2.5 ng/ml)- stimulated glycogenolysis. Isoproterenol (1 micro gram/ml) stimulated phosphorylase b to a conversion in hemidiaphragms of both strains to give a phosphorylase a (minus AMP) reactivity of 1.6 micro units/gram in I strain and 2.2 micro units/gram in C57BL. At this isoproterenol concentration the glycogenolytic response in I strain hemidiaphragms was 76% of that of C57BL with a phosphorylase a concentration of 73% of that of C57BL. The correlation of the rate of glycogenolysis and the phosphorylase a concentration in I strain hemidiaphragms suggests that in mutant skeletal muscle, as in that of control strains, phosphorylase a is the primary catalyst of catecholamine-stimulated glycogenolysis.